1. Simple ELISA protocol 1. Coat antigen onto microplate
2. Allow protein adsorption and block unoccupied sites with neutral protein
3. Add antibody solution into each well
4. Add HRP or AP conjugated secondary antibody into each well and develop colorimetric reaction with appropriate substrate
5. Read absorbance in spectrophotometer with appropriate filter and quantitate relative antigen levels

2. ELISA (or Western Blot)
Antigen + antibody bind – wash
Antigen + antibody + anti-antibody-enzyme – wash
Antigen + antibody + anti-antibody- enzyme + substrate
Determine (quantitate) color reaction

3. Home pregnancy test

4. ELISA
Direct
Indirect

5. Direct and indirect immunofluorescence

6. Western Blot

7. Western blot Look at Western Blot Protein electrophoresis
Staining protein gel

8. Protein electrophoresis

9. To determine size

10. Other protein techniques

11. 2D electrophoresis – 2 steps
Take the isoelectric focusing gel (IEF)
Proteins separated based on charge
Place it on top of the SDS gel
Separated based on size
Run gel – proteins run from the IEF gel into the SDS gel
Stain gel – analyze different spots

12. 2D-electrophoresis

13. Picture of a 2D gel